Optimising catalytic properties of supported sulfonic acid catalysts

Siliceous mesoporous molecular sieves (SBA-15) have been functionalised with propylsulfonic acid groups by both co-condensing 3-mercaptopropyltrimethoxysilane with the solid at the synthesis (sol-gel) stage and by grafting the same compound to pre-prepared SBA-15, followed, in both cases, by oxidation to sulfonic acid. The acidic and catalytic properties of the supported sulfonic acids prepared in the two ways have been compared, using ammonia adsorption calorimetry and the benzylation reaction between benzyl alcohol and toluene. Using a combination of X-ray photoelectron spectroscopy and other analytical techniques, the level of functionalisation and the extent of subsequent oxidation of tethered thiol to sulfonic acid, both in the bulk and close to the surface of SBA-15 particles, have been assessed. The research shows that the co-condensing route leads to higher levels of functionalisation than the grafting route. The extent of oxidation of added thiol to acid groups is similar using the two routes, about 70% near the surface and only 50% in the bulk. Comparison is made with polymer supported sulfonic acid catalysts, Amberlysts 15 and 35, and Nafion. Nafion shows the highest acid strength and the highest specific catalytic activity of all materials studied. Amongst the other materials, average acid strengths are broadly similar but there appears to be a relationship between the concentration of acid sites on the catalysts and their specific activity in the benzylation reaction. A model is proposed to explain this, in which clustering of sulfonic acid groups, even to a small extent, leads to disproportionately enhanced catalytic activity. © 2009 Elsevier B.V. All rights reserved.

Effect of nitric oxide on apoptotic activity in the rat gastrointestinal tract

The effect of nitric oxide (NO) on apoptosis in the gastrointestinal mucosa was investigated. Experiments involved long-term exposure of rat gastric mucosal cells in vitro to exogenous NO delivered from the NO, donor S-nitroso-N-acetyl-penicillamine, and the effect of intravenous administration of lipopolysaccharide in vivo, in the presence and absence of the selective inhibitor of inducible NO synthase N-(3-(aminomethyl)benzyl) acetamidine (1400 W). S-nitroso-N-acetyl-penicillamine produced a dose-related inhibition of caspase 3-like activity and DNA fragmentation in isolated gastric mucosal cells. Caspase 3-like activity and DNA fragmentation in gastric, ileal and colonic mucosa were increased both 5 and 24 h after injection of lipopolysaccharide (3 mg/kg, i.v.) to rats in vivo. Administration of 1400 W (5 mg/kg, i.v.) immediately after lipopolysaccharide enhanced caspase 3-like activity and DNA fragmentation above that found with lipopolysaccharide alone. In conclusion, data obtained both in vitro and in vivo suggest that NO exerts an anti-apoptotic effect on rat gastrointestinal mucosal cells. © 2001 Elsevier Science B.V.

Mesoporous sulfonic acid silicas for pyrolysis bio-oil upgrading via acetic acid esterification

Propylsulfonic acid derivatised SBA-15 catalysts have been prepared by post modification of SBA-15 with mercaptopropyltrimethoxysilane (MPTMS) for the upgrading of a model pyrolysis bio-oil via acetic acid esterification with benzyl alcohol in toluene. Acetic acid conversion and the rate of benzyl acetate production was proportional to the PrSO3H surface coverage, reaching a maximum for a saturation adlayer. Turnover frequencies for esterification increase with sulfonic acid surface density, suggesting a cooperative effect of adjacent PrSO3H groups. Maximal acetic acid conversion was attained under acid-rich conditions with aromatic alcohols, outperforming Amberlyst or USY zeolites, with additional excellent water tolerance.

Antimicrobial effect of Artemisia species on Pseudomonas aeruginosa

Antibiotic resistance has emerged as a severe problem in hospital-acquired infectious disease. The Gram-negative bacterium Pseudomonas aeruginosa is found to cause secondary infection in immune-compromised patients. Unfortunately, it is resistant to virtually all β-lactam antibiotics such as penicillin, cephalosporin and others. Researchers are seeking for new compounds to treat several antibiotic-resistant bacterial strains. Artemisia plant extracts are commonly used for their therapeutic properties by natives throughout dry regions of North and South America. Here, they are administered as an alternative medicine for stomach problems and other complex health issues. In this study, the antimicrobial effects of plant extracts from several Artemisia species as well as compounds dehydroleucodine and dehydroparishin-B (sesquiterpenes derived specifically from A. douglasiana) were used as treatments against the pathogenicity effects of P. aeruginosa. Results showed that both compounds effectively inhibit the secretion of LasB elastase, biofilm formation and type III secretion, but fail to control LasA protease. This is a significant observation because these virulent factors are crucial in establishing P.aeruginosa infection. The results from this study signify a plausible role for future alternative therapy in the biomedical field, which recommends DhL and DhP can be studied as key compounds against bacterial infections of Pseudomonas aeruginosa.

Asymmetric syntheses of analogs of kainic acid

Kainic acid has been used for nearly 50 years as a tool in neuroscience due to its pronounced neuroexcitatory properties. However, the significant price increase of kainic acid resulting from the disruption in the supply from its natural source, the alga Digenea Simplex, as well as inefficient synthesis of kainic acid, call for the exploration of functional mimics of kainic acid that can be synthesized in a simpler way. Aza kainoids analog could be one of them. The unsubstituted aza analog of kainoids has demonstrates its ability as an ionotropic glutamate receptor agonist and showed affinity in the chloride dependent glutamate (GluCl) binding site. This opened a question of the importance of the presence of one nitrogen or both nitrogens in the aza kainoid analogs for binding to glutamate receptors. Therefore, two different pyrrolidine analogs of kainic acid, trans -4-(carboxymethyl)pyrrolidine-3-carboxylic acid and trans -2-carboxy-3-pyrrolidineacetic acid, were synthesized through multi-step sequences. The lack of the affinity of both pyrrolidine analogs in GluCl binding site indicated that both nitrogens in aza kainoid analogs are involved in hydrogen bonding with receptors, significantly enhancing their affinity in GluCl binding site. Another potential functional mimic of kainic acid is isoxazolidine analogs of kainoids whose skeleton can be constituted directly via a 1, 3 dipolar cycloaddition as the key step. The difficulty in synthesizing N-unsubstituted isoxazolidines when applying such common protecting groups as alkyl, phenyl and benzyl groups, and the requirement of a desired enantioselectivity due to the three chiral ceneters in kainic acid, pose great challenges. Hence, several different protected nitrones were studied to establish that diphenylmethine nitrone may be a good candidate as the dipole in that the generated isoxazolidines can be deprotected in mild conditions with high yields. Our investigations also indicated that the exo/endo selectivity of the 1, 3 dipolar cycloaddition can be controlled by Lewis acids, and that the application of a directing group in dipolarophiles can accomplish a satisfied enantioselectivity. Those results demonstrated the synthesis of isoxazoldines analogs of kainic acid is very promising.

Diversity of Vancomycin Genes in Sub-tropical Soils

With the increased antibiotic exposure from anthropogenic sources, soil microbes are an ever-increasing ecological pool of resistant bacteria. This is the case with bacterial resistance to vancomycin through transfer of van-resistance genes by transposons. Studies show that bacterial species other than enteroccoci harbor genetic-like elements such as the Tn1546 transposon containing vancomycin-resistant genes. Overuse and misuse of antibiotics in hospital settings and agricultural practices have led to an increase in transferability of vancomycin-resistant genes among microbes. The objective of this project is to analyze the diversity of these genes found in the soil microbes from Miami-Dade County. Bacterial isolates were Gram-stained and the Kirby-Bauer antibiotic disk diffusion test was performed to determine the degree of resistance. Results showed that all bacterial isolates were resistant to penicillin at the 10 µg concentration and most were susceptible to varying vancomycin concentrations (10 µg, 20 µg, and 30 µg). A 1465 bp fragment was amplified from the 16S rDNA gene using 27F and 1492R universal primers from the multi-antibiotic resistant bacteria and sequenced to identify the isolates. Three Gram-negative bacteria genera were identified with the closest phylogenetic match to: Pseudomonas sp., Stenotrophomonas sp., Xanthomonas sp., as well as two Gram-positive bacteria genera: Bacillus sp. and Brevibacillus sp. The isolates’ vanA and vanB genes were amplified using the respective primers. Ongoing work is underway to sequence and compare these known van resistant genes, with the goal of revealing intrinsic vancomycin resistance present in soil bacteria.

Frequência de microrganismos em conteúdo vulvovaginal de vacas tratadas com dispositivos intravaginais de progesterona

The dissertation was divided in two studies. With the first, aimed to evaluate the occurrence of microorganisms present in the vulvovaginal region of cows that received intravaginal progesterone devices during the fixed-time artificial insemination (FTAI) programs, and correlate the results with pregnancy rates. Samples were collected from vulvovaginal region of 30 beef cows Guzerá and 30 crossbred dairy cows, and also intravaginal devices, randomly. Of the 120 samples of cows, 60 corresponded to the collections of the period prior to the introduction device (D0) and 60 to the subsequent withdrawal of it (D9); it yielded 100% of bacterial growth, whereas, in most samples, it was found more than one isolated. In D0, the most frequent agent was Escherichia coli (52%), and in D9, Proteus spp and E. coli were the most frequent (32% and 28%, respectively). Regarding intravaginal progesterone devices, in D0 were isolated 37 microorganisms, being predominant those of the genus Bacillus (35%); in D9, 41 colony forming units (CFU) were isolated, of which 36.6% corresponded to Proteus spp. For the analysis of the antimicrobial profile, susceptibility testing was performed by diffusion agar disk, and cows that did not became pregnant after FTAI program were selected, as a future treatment. There resistance 100% to penicillin, and sensitivity, approximately, 90% to gentamicin, both isolates obtained from samples of beef cows and obtained of dairy cows. Regarding pregnancy rate, the 30 beef cows, 11 were diagnosed pregnant (36.7%), 4 (36.4%) treated with reused devices and 7 (63.6%) with new devices, which showed more effective. Of the 30 dairy cows, 15 were pregnant (50%), 8 (53.3%) were implanted with reused devices and 7 (46.7%) with new devices, with no significant differences in pregnancy rates. Because it is a research, females were chosen at random, and factors such as body condition, nutritional management and health weren't priority. With the second study, aimed to analyze the similarity between strains, conducted by technical Random Amplified Polymorphic DNA (RAPD -PCR). Presence of E. coli and the absence of pregnancy were selection criteria used. From the results, it was observed that most of the isolates wasn't phylogenetically similar, since they showed lower than 85% similarity. The study stressed the importance of E. coli in vulvovaginal microbiota of cows and the presence of phenotypic and genotypic characters of this bacterium on possible reproductive problems.

Studies in asymmetric and heterocyclic synthesis: I. chiral ketones II. Quinolones III. Trifluoromethylated pyrones

This thesis is split into three sections based on three different areas of research. In the first section, investigations into the α-alkylation of ketones using a novel chiral auxiliary is reported. This chiral auxiliary was synthesised containing a pyrrolidine ring in the chiral arm and was applied in the preparation of α-alkylated ketones which were obtained in up to 92% ee and up to 63% yield over two steps. Both 3-pentanone and propiophenone based ketones were used in the investigation with a variety of both alkyl and benzyl based electrophiles. The novel chiral auxiliary was also successful when applied to Michael and aldol reactions. A diamine precursor en route to the chiral auxiliary was also applied as an organocatalyst in a Michael reaction, with the product obtained in excellent enantioselectivity. In the second section, investigations into potential anti-quorum sensing molecules are reported. The bacteria Pseudomonas aeruginosa is an antibiotic-resistant pathogen that demonstrates cooperative behaviours and communicates using small chemical molecules in a process termed quorum sensing. A variety of C-3 analogues of the quorum sensing molecules used by P. aeruginosa were synthesised. Expanding upon previous research within the group, investigations were carried out into alternative protecting group strategies of 2-heptyl-4-(1H)- quinolone with the aim of improving the yields of products of cross-coupling reactions. In the third section, investigations into fluorination and trifluoromethylation of 2-pyrones, pyridones and quinolones is reported. The incorporation of a fluorine atom or a trifluoromethyl group into a molecule is important in pharmaceutical drug discovery programmes as it can lead to increased lipophilicity and bioavailability, however late-stage incorporation is rarely reported. Both direct fluorination and trifluoromethylation were attempted. Eight trifluoromethylated 2-pyrones, five trifluoromethylated 2-pyridones and a trifluoromethylated 2-quinolone were obtained in a late-stage synthesis from their respective iodinated precursors using methyl fluorosulfonyldifluoroacetate as a trifluoromethylating reagent.

Asymmetric oxidation of sulfides

This review discusses synthesis of enantiopure sulfoxides through the asymmetric oxidation of prochiral sulfides. The use of metal complexes to promote asymmetric sulfoxidation is described in detail, with a particular emphasis on the synthesis of biologically active sulfoxides. The use of non-metal-based systems, such as oxaziridines, chiral hydroperoxides and peracids, as well as enzyme-catalyzed sulfoxidations is also examined.

Molecular characterization of extended-spectrum cephalosporin-resistance in Escherichia coli in pigs on-farm and from clinical cases throughout Quebec, Canada during 16 years

Le développement de la multirésistance chez Escherichia coli est un problème important en médecine animale et humaine. En outre, l’émergence et la diffusion des déterminants de résistance aux céphalosporines à larges spectres de troisième génération (ESCs) parmi les isolats, incluant des céphalosporines essentielles en médecine humaine (ex. ceftriaxone et ceftiofur), est un problème majeur de santé publique. Cette thèse visait trois objectifs. D’abord étudier la dynamique de la résistance aux antimicrobiens (AMR) ainsi que la virulence et les profils génétiques de la AMR des E. coli isolées de porcs recevant une nourriture post-sevrage supplémentée avec de la chlortétracycline et de la pénicilline G, et, accessoirement, évaluer les effets d'additifs alimentaires sur cette dynamique en prenant pour exemple d'étude un minéral argileux, la clinoptilolite, étant donné son possible lien avec le gène blaCMY-2 qui confère la résistance au ceftiofur. L'objectif suivant était d'investiguer les mécanismes menant à une augmentation de la prévalence du gène blaCMY-2 chez les porcs qui reçoivent de la nourriture médicamentée et qui n'ont pas été exposés au ceftiofur Ici encore,nous avons examiné les effets d’un supplément alimentaire avec un minéral argileux sur ce phénomène. Enfin, notre dernier objectif était d’étudier, dans le temps, les génotypes des isolats cliniques d'E. coli résistant au ceftiofur, isolés de porcs malades au Québec à partir du moment où la résistance au ceftiofur a été rapportée, soit de 1997 jusqu'à 2012. Dans l'étude initiale, la prévalence de la résistance à 10 agents antimicrobiens, incluant le ceftiofur, s’accroît avec le temps chez les E.coli isolées de porcelets sevrés. Une augmentation tardive de la fréquence du gène blaCMY-2, encodant pour la résistance au ceftiofur, et la présence des gènes de virulence iucD et tsh a été observée chez les isolats. La nourriture supplémentée avec de la clinoptilolite a été associée à une augmentation rapide mais, par la suite, à une diminution de la fréquence des gènes blaCMY-2 dans les isolats. En parallèle, une augmentation tardive dans la fréquence des gènes blaCMY-2 et des gènes de virulence iucD et tsh a été observée dans les isolats des porcs contrôles, étant significativement plus élevé que dans les porcs ayant reçu l'additif au jour 28. La diversité, au sein des E. coli positives pour blaCMY-2 , a été observée au regard des profils AMR. Certaines lignées clonales d'E.coli sont devenues prédominantes avec le temps. La lignée clonale du phylotype A prédominait dans le groupe supplémenté, alors que les lignées clonales du phylotype B1, qui possèdent souvent le gène de virulence iucD associé aux ExPEC, prédominaient dans le groupe contrôle. Les plasmides d'incompatibilité (Inc) des groupes, I1, A/C, et ColE, porteurs de blaCMY-2, ont été observés dans les transformants. Parmi les souches cliniques d'E.coli ESC-résistantes, isolées de porcs malades au Québec de 1997 à 2012, blaCMY-2 était le gène codant pour une β-lactamase le plus fréquemment détecté; suivi par blaTEM et blaCTX-M,. De plus, les analyses clonales montrent une grande diversité génétique. Par contre, des isolats d'E. coli avec des profils PFGE identiques ont été retrouvés dans de multiples fermes la même année mais aussi dans des années différentes. La résistance à la gentamicine, kanamycine, chloramphenicol, et la fréquence de blaTEM et de IncA/C diminuent significativement au cour de la période étudiée, alors que la fréquence de IncI1 et de la multirésistance à sept catégories d'agents antimicrobiens augmente significativement avec le temps. L'émergence d'isolats d'E. coli positifs pour blaCTX-M, une β-lactamase à large spectre et produisant des ESBL, a été observée en 2011 et 2012 à partir de lignées clonales distinctes et chez de nombreuses fermes. Ces résultats, mis ensemble, apportent des précisions sur la dissémination de la résistance au ceftiofur dans les E. coli isolées de porcs. Au sein des échantillons prélevés chez les porcs sevrés recevant l'alimentation médicamentée sur une ferme, et pour laquelle une augmentation de la résistance au ceftiofur a été observée, les données révèlent que les souches d'E. coli positives pour blaCMY-2 et résistantes aux ESCs appartenaient à plusieurs lignées clonales différentes arborant divers profils AMR. Le gène blaCMY-2 se répand à la fois horizontalement et clonalement chez ces E. coli. L'ajout de clinoptilotite à la nourriture et le temps après le sevrage influencent la clonalité et la prévalence du gène blaCMY-2 dans les E. coli. Durant les 16 années d'étude, plusieurs lignées clonales différentes ont été observées parmi les souches d'E. coli résistantes au ceftiofur isolées de porc malades de fermes québécoises, bien qu’aucune lignée n'était persistante ou prédominante pendant l'étude. Les résultats suggèrent aussi que le gène blaCMY-2 s'est répandu à la fois horizontalement et clonalement au sein des fermes. De plus, blaCMY-2 est le gène majeur des β-lactamases chez ces isolats. À partir de 2011, nous rapportons l'émergence du gène blaCTX-M dans des lignées génétiques distinctes.

Employment of the Technique of Radiotracers for Analysis of Industrial Filters

The main aim of this work is to develop a methodology to evaluate the characteristics of porous media in filter using the radio-tracing technique. To do this, an experimental prototype filter made up of an acrylic cylinder, vertically mounted and supported on the lower side by a controlled leaking valve was developed. Two filters (spheres of acrylic and silica crystals) were used to check the movement of the water through the porous media using 123I in its MIBG (iodine-123-meta-iodo benzyl-guanidine) form. Further up the filter an instantaneous injection of the substance makes it possible to see the passage of radioactive clouds through the two scintillatory detectors NaI (2x2)” positioned before and immediately after the cylinder with the filtering element (porous media). The are caused by the detectors on the passage of the radioactive cloud are analyzed through statistical functions using the weighted moment method which makes it possible to calculate the Residence-Time (the amount of time the tracer takes to thoroughly pass through the filter) per the equation of dispersion in tubular flow and the one-directional flow of the radiotracer in the porous media.

As doenças infeciosas e a história dos antibióticos

O presente trabalho aborda a história das principais doenças que assolaram a Europa desde a época medieval à época contemporânea, incluindo a referência às origens, às manifestações e ao tratamento. É igualmente abordada a questão do isolamento das substâncias ativas nas plantas medicinais e farmacológicas até à descoberta do primeiro antibiótico identificado pelo homem – a penicilina, por Alexandre Fleming – e a sua enorme importância no combate aos micro-organismos.

Degradation of Biofumigant Isothiocyanates and Allyl Glucosinolate in Soil and Their Effects on the Microbial Community Composition

Brassicales species rich in glucosinolates are used for biofumigation, a process based on releasing enzymatically toxic isothiocyanates into the soil. These hydrolysis products are volatile and often reactive compounds. Moreover, glucosinolates can be degraded also without the presence of the hydrolytic enzyme myrosinase which might contribute to bioactive effects. Thus, in the present study the stability of Brassicaceae plant-derived and pure glucosinolates hydrolysis products was studied using three different soils ( model biofumigation). In addition, the degradation of pure 2-propenyl glucosinolate was investigated with special regard to the formation of volatile breakdown products. Finally, the influence of pure glucosinolate degradation on the bacterial community composition was evaluated using denaturing gradient gel electrophoresis of 16S rRNA gene amplified from total community DNA. The model biofumigation study revealed that the structure of the hydrolysis products had a significant impact on their stability in the soil but not the soil type. Following the degradation of pure 2-propenyl glucosinolate in the soils, the nitrile as well as the isothiocyanate can be the main degradation products, depending on the soil type. Furthermore, the degradation was shown to be both chemically as well as biologically mediated as autoclaving reduced degradation. The nitrile was the major product of the chemical degradation and its formation increased with iron content of the soil. Additionally, the bacterial community composition was significantly affected by adding pure 2-propenyl glucosinolate, the effect being more pronounced than in treatments with myrosinase added to the glucosinolate. Therefore, glucosinolates can have a greater effect on soil bacterial community composition than their hydrolysis products.

Functional studies of lignin biosynthesis genes and putative flowering gene in Miscanthus x giganteus and studies on indolyl glucosinolate biosynthesis and translocation in Brassica oleracea

The first topic area of this thesis involved studies on the accumulation and translocation of glucosinolates (GSs), bioactive secondary plant compounds, in broccoli plants. Changes in GS accumulation and gene expression levels in response to exogeneous methyl jasmonate (MeJA) treatment were analyzed in different tissue types at different developmental stages of broccoli. Greater accumulation of GSs with MeJA treatment was observed in apical leaves of broccoli seedlings and florets of plants at harvest maturity. Increases in indolyl GS in apical leaves of seedlings and florets were coupled with the up-regulation of indolyl GS biosynthesis genes. The accumulation of indolyl GSs appears to be modulated by MeJA treatment in an organ-specific manner for optimal distribution of defense substances in the plant. Metabolic profiling of hydrophilic metabolites using GC-MS demonstrated increased accumulation of various phenolics, ascorbates and amino acids in broccoli tissues after MeJA treatment. Distinct changes in carbohydrate levels observed between different tissues (vegetative leaves and floret tissues) of broccoli plants after treatment suggest that carbon metabolism is differentially modulated by MeJA treatment in different tissue types depending on sink-source relationships. Reduced levels of hexose sugars and tricarboxylic acid intermediates after MeJA treatment may reflect the increased requirement for carbon and energy needed to drive secondary product biosynthesis to accumulate metabolites for defense against insects and other herbivores. Substantial increases of indolyl and aromatic GSs after exogenous treatment with MeJA in stem and petioles of seedlings and the existence of intact indolyl-GS forms in phloem exudates suggest enhanced de novo synthesis in combination with active transport. Indoly GSs share structural similarities with the auxin, IAA, and may interact with components of the auxin transport system for intra- and extra-cellular transport or translocation. Application of the auxin efflux inhibitor, 1-naphthylphthalamic acid (NPA) reduced MeJA-mediated accumulation of indolyl GSs in broccoli florets and seedling tissues. NPA did not inhibit expression of indolyl GS biosynthesis genes shown to be upregulated by MeJA treatment or the accumulation of tryptophan, the amino acid precursor of indolyl GSs. Exogenous application of benzyl GS to Arabidopsis roots induced ectopic expression of the PIN1 protein associated with the auxin transport system similar to treatment with NPA, again suggesting GS interaction with the auxin efflux carrier system. The inhibitory effect of NPA on MeJA-mediated accumulation of GS may be due to competitive binding of NPA to auxin efflux carrier components and that GS transport is mediated by the auxin transport system. The inhibitory effect of NPA on indolyl and aromatic GS accumulation and the bioactivity of exogenous treatment of these GS compounds in PIN1 localization, Arabidopsis root growth, and gravitrophic response suggest that indolyl and aromatic GSs may be antagonistic to IAA transport and biosynthesis. Indolyl and aromatic GSs can also be potentially converted into IAA by hydrolysis. This intrinsic feature of GSs may be the part of a sophisticated regulatory process where the metabolic pathways in the plant shift from active growth to a reversible defense posture in response to biotic or abiotic stress. It seems likely that indolyl and aromatic GSs are important compounds that provide connections between jasmonate and auxin signaling. Further studies are required to reveal the regulatory mechanism for crosstalk between the two hormones. The third part of this research was to investigate effect of selenium fertilization and MeJA treatment on accumulation of GSs in broccoli florets. Increasing dietary intake of the element selenium (Se) has been shown to reduce the risk of cancer. Simultaneous enhancement of both Se and GS concentrations in broccoli floret tissue were conducted through the combined treatment of MeJA with Se fertilization. A low level of Se fertilization (concentration) with MeJA treatment displayed no significant changes in total aliphatic GS concentrations with 90% and 50% increases in indolyl and total GSs concentrations, respectively. This result suggests that Se- and GS-enriched broccoli with improved health-promoting properties can be generated by this combined treatment. The second topic of this thesis was conducted to provide basic information required to improve biomass quality and productivity and develop tools for gene transformation in Miscanthus x giganteus. The perennial rhizomatous grass, Miscanthus x giganteus is an ideal biomass crop due to its rapid vegetative growth and high biomass yield potential. As a naturally occurring sterile hybrid, M. x giganteus must be propagated vegetatively by mechanicalling divided rhizomes or from micropropagated plantlets. The effect of callus type, age and culture methods on regeneration competence was studied to improve regeneration efficiency and shorten the period of tissue culture in M. x giganteus propagation. Seven lignin biosynthesis genes and one putative flowering gene were isolated from M. x giganteus by PCR reactions using maize othologous sequences. Southern hybridization and nuclear DNA content analysis indicated that the genes isolated from M. x giganteus exist in the genome of other Miscanthus species as multiple copies. Analysis of lignin content and histological staining of lignin deposition indicated that higher lignin content is found in mature stem node tissues compared to young leaves and apical stem nodal tissues. Cell wall lignification is associated with increasing tissue maturity in Miscanthus species. RNAi and antisense constructs harboring sequences of these genes were developed to generate Miscanthus transgenic plants with suppressed of lignin biosynthesis and delayed flowering.